JS-K, a glutathione/glutathione S-transferase-activated nitric oxide releasing prodrug inhibits androgen receptor and WNT-signaling in prostate cancer cells.

BACKGROUND: Nitric oxide (NO) and its oxidative reaction products have been repeatedly shown to block steroid receptor function via nitrosation of zinc finger structures in the DNA-binding domain (DBD). In consequence NO-donors could be of special interest for the treatment of deregulated androgen receptor(AR)-signaling in castration resistant prostate cancer (CRPC). METHODS: Prostate cancer (PCa) cells were treated with JS-K, a diazeniumdiolate derivate capable of generating large amounts of intracellular NO following activation by glutathione S-transferase. Generation of NO was determined indirectly by the detection of nitrate in tissue culture medium or by immunodetection of nitrotyrosine in the cytoplasm. Effects of JS-K on intracellular AR-levels were determined by western blotting. AR-dimerization was analyzed by mammalian two hybrid assay, nuclear translocation of the AR was visualized in PCa cells transfected with a green fluorescent AR-Eos fusion protein using fluorescence microscopy. Modulation of AR- and WNT-signalling by JS-K was investigated using reporter gene assays. Tumor cell proliferation following JS-K treatment was measured by MTT-Assay. RESULTS: The NO-releasing compound JS-K was shown to inhibit AR-mediated reporter gene activity in 22Rv1 CRPC cells. Inhibition of AR signaling was neither due to an inhibition of nuclear import nor to a reduction in AR-dimerization. In contrast to previously tested NO-donors, JS-K was able to reduce the intracellular concentration of functional AR. This could be attributed to the generation of extremely high intracellular levels of the free radical NO as demonstrated indirectly by high levels of nitrotyrosine in JS-K treated cells. Moreover, JS-K diminished WNT-signaling in AR-positive 22Rv1 cells. In line with these observations, castration resistant 22Rv1 cells were found to be more susceptible to the growth inhibitory effects of JS-K than the androgen dependent LNCaP which do not exhibit an active WNT-signaling pathway. CONCLUSIONS: Our results suggest that small molecules able to inhibit WNT- and AR-signaling via NO-release represent a promising platform for the development of new compounds for the treatment of CRPC.

AR-Q640X, a model to study the effects of constitutively active C-terminally truncated AR variants in prostate cancer cells.

PURPOSE: A recently identified mechanism allowing prostate cancer (PCa) cells to grow in the absence of androgens is the expression of constitutively active, C-terminally truncated androgen receptor (AR) variants lacking vast parts of the ligand-binding domain. These AR variants termed ARΔLBD are either products of alternative splicing, point mutations leading to premature stop codons or proteolytic cleavage of the AR. Some controversies exist about the requirement of additional full-length AR for the full transcriptional activity of the ARΔLBD. On basis of a mutated, C-terminally truncated AR termed Q640X, we developed an experimental model for the study of ARΔLBD in PCa cells. METHODS: Activation of AR-dependent promoters was analyzed by reporter gene assays. Dimerization studies were conducted using a mammalian two-hybrid system. RESULTS: Although Q640X/Q640X homodimers were able to induce the expression of certain AR target genes, Q640X/AR heterodimers were necessary to activate the full panel of androgen-dependent genes under androgen-deprived conditions. CONCLUSIONS: The following study supports the hypothesis that castration-resistant prostate cancer (CRPC) cells are able to activate specific androgen-dependent genes by selective modulation of the ratio between ARΔLBD and their putative dimerization partners like the full-length AR or other ARΔLBD in the absence of androgens. The present data suggest that AR-mutant Q640X is a powerful experimental tool for the functional analysis of ARΔLBD in CRPC.

Effects of sorafenib on C-terminally truncated androgen receptor variants in human prostate cancer cells.

Recent evidence suggests that the development of castration resistant prostate cancer (CRPCa) is commonly associated with an aberrant, ligand-independent activation of the androgen receptor (AR). A putative mechanism allowing prostate cancer (PCa) cells to grow under low levels of androgens, is the expression of constitutively active, C-terminally truncated AR lacking the AR-ligand binding domain (LBD). Due to the absence of a LBD, these receptors, termed ARΔLBD, are unable to respond to any form of anti-hormonal therapies. In this study we demonstrate that the multikinase inhibitor sorafenib inhibits AR as well as ARΔLBD-signalling in CRPCa cells. This inhibition was paralleled by proteasomal degradation of the AR- and ARΔLBD-molecules. In line with these observations, maximal antiproliferative effects of sorafenib were achieved in AR and ARΔLBD-positive PCa cells. The present findings warrant further investigations on sorafenib as an option for the treatment of advanced AR-positive PCa.

Experimental and analytical datasets for pressure drop and flow distribution in a Z-type flat-plate solar absorber.

This data article provides two dataset types for total pressure drop and parallel-flow distribution in a Z-type flat-plate solar thermal absorber with water as the working fluid. The first dataset consists of high-resolution pressure drop measurements at different temperatures under laminar and turbulent flow conditions obtained experimentally using a state-of-the-art hydraulic test rig. The second dataset comprises analytical data on flow distribution in the absorber. Conducting high-resolution pressure measurements, essential for evaluating thermo-hydraulic models, is a sensitive and time-demanding process requiring a relatively elaborated test rig to accurately measure pressure drop at different temperatures and flow rates in the presence of thermal equilibrium. In this context, engineers and researchers can use these datasets to compare and verify developed numerical models for thermo-hydraulic evaluation of pressure drop and flow distribution in flat-plate solar collectors under both laminar and turbulent flow regimes. The article also comprises analytical data for flow distribution in the absorber for several header configurations presented by dimensionless-flow-rate and non-uniformity. This data article is related to the research article (Shantia et al., 2022 ). The datasets are accessible in the supplementary files accompanied by the online version of this article and in the Mendeley Data repository.

Stilbene induced inhibition of androgen receptor dimerization: implications for AR and ARΔLBD-signalling in human prostate cancer cells.

BACKGROUND: Advanced castration resistant prostate cancer (CRPC) is often characterized by an increase of C-terminally truncated, constitutively active androgen receptor (AR) variants. Due to the absence of a ligand binding domain located in the AR-C-terminus, these receptor variants (also termed ARΔLBD) are unable to respond to all classical forms of endocrine treatments like surgical/chemical castration and/or application of anti-androgens. METHODOLOGY: In this study we tested the effects of the naturally occurring stilbene resveratrol (RSV) and (E)-4-(2, 6-Difluorostyryl)-N, N-dimethylaniline, a fluorinated dialkylaminostilbene (FIDAS) on AR- and ARΔLBD in prostate cancer cells. The ability of the compounds to modulate transcriptional activity of AR and the ARΔLBD-variant Q640X was shown by reporter gene assays. Expression of endogenous AR and ARΔLBD mRNA and protein levels were determined by qRT-PCR and Western Blot. Nuclear translocation of AR-molecules was analyzed by fluorescence microscopy. AR and ARΔLBD/Q640X homo-/heterodimer formation was assessed by mammalian two hybrid assays. Biological activity of both compounds in vivo was demonstrated using a chick chorioallantoic membrane xenograft assay. RESULTS: The stilbenes RSV and FIDAS were able to significantly diminish AR and Q640X-signalling. Successful inhibition of the Q640X suggests that RSV and FIDAS are not interfering with the AR-ligand binding domain like all currently available anti-hormonal drugs. Repression of AR and Q640X-signalling by RSV and FIDAS in prostate cancer cells was caused by an inhibition of the AR and/or Q640X-dimerization. Although systemic bioavailability of both stilbenes is very low, both compounds were also able to downregulate tumor growth and AR-signalling in vivo. CONCLUSION: RSV and FIDAS are able to inhibit the dimerization of AR and ARΔLBD molecules suggesting that stilbenes might serve as lead compounds for a novel generation of AR-inhibitors.